Anti cd11b pe clone m1 70

Peritoneal cells from Scn8a^dmu/+ and Scn8a^+/+ mice (n = 16 each) were harvested by i.p. injection and recovery of 4 ml PBS, 0.5% bovine serum albumin, 5 mM EDTA. Mouse peritoneal cavity cells (PCC) were counted. Cells were centrifuged at 400 x g for 5 minutes at 4°C and resuspended in a 1/1000 dilution of fixable viability dye (FVD) eFluor 450 (eBiosciences) in PBS for 20 minutes. Then the cells were washed with wash buffer (PBS + 2% FBS + 20 mM NaN₃), centrifuged and resuspended in antibody cocktails containing anti-CD19-BV510 (clone 6D5, BioLegend), anti-CD11b-PE-eFL610 (clone M1/70, eBiosciences), CD117 (c‐kit)‐PE (clone 2B8, BioLegend), anti-CD4-APC (clone GK1.5, eBiosciences), anti-FcϵRI-APC eFL780 (clone Mar-1, eBioscience), anti-CD8a-BV650 (clone 53-6.7, BD Biosciences), anti-CD3-BV711 (clone 145-2C11, BD Biosciences), anti-Siglec-F-PE-CF594 (clone E50-2440, BD Bioscience), anti-CD11b-PE (clone M1/70, eBiosciences), anti-F4/80-PE-Cy7 (clone BM8, eBioscience), anti-Ly6C-APC-eF780 (clone HK1.4, eBioscience), or anti-Ly6G-BV605 (clone 1A8, BD Biosciences) for 30 minutes. Stained fixed cells were acquired for analysis using a BD LSRFortessa and results were analyzed using FlowJo (Ashland, OR) software. A visual representation of the gating strategy used to identify the cell populations in the mouse peritoneum is provided (Supplementary Figure 2).

Staining for cellular and nuclear morphology was performed on cytospins from cultures of progenitors or differentiated macrophages by incubating with Giemsa solution (Merck, Darmstadt, Germany) after methanol fixation. Analysis by brightfield microscopy was performed using a Keyence BZ9000 microscope at a magnification of 20x (Keyence, Neu-Isenburg, Germany).
Expression of cell surface markers was measured after blocking of unspecific binding sites with CD16/CD32 Fc-Block (BD Biosciences, San Jose, CA, USA). Cells were stained with anti-CD11b-PE (clone M1/70, #12-0112, eBioscience, San Diego, CA, USA), anti-CD11c-PE (clone HL3, #553802, BD), or anti-F4/80-AF647 (clone A3-1, #MCA497A647, Biolegend, San Diego, CA, USA) followed by flow cytometry analysis on a FACS Calibur (BD Biosciences, Heidelberg, Germany).